The mouse ovary serves as a paradigm for investigating the developmental biology of mammalian gonadogenesis, oogenesis, and fertilization. Gonadogenesis: Mouse gestation occurs over 20 days. Germ cells, first detected in the developing embryo 7.5 days post coitus (dpc), migrate from the allantois tot he genital ridge by 12.5 dpc. In XX females, the primitive gonad then differentiates into an ovary. Poly(A+) was purified from female and male genital ridges isolated 12-13 dpc. cDNA libraries have been made for each sex and substractive cloning techniques are being used to identify female-specific gene products involved in early sex determination and gonadogenesis. Oogenesis: The zona pellucida, composed of three glycoproteins (ZP1, ZP2, ZP3), is an ovary-specific extracellular matrix that surrounds growing oocytes. We have cloned the three mouse and human genes and have demonstrated in mice that their coordinate expression is restricted to oocytes. Approximately 200bp upstream of the start of transcription of each of the three genes is an E-box that is necessary and sufficient for high level reporter gene expression in oocytes. By gel mobility shift assays, we have identified a heterodimer composed of basic helix-loop- helix transcription factors (the ubiquitously expressed E12 and ZAP-1, Zona Activating Protein) that bind to this E-box. These studies provide insight into the molecular basis for oocyte-specific gene expression and identify early markers of oocyte growth and differentiation. Fertilization: The three zona proteins are secreted and form an extracellular matrix that mediates the relatively species-species- specific events of fertilization. To assess protein-protein interactions in the assemblage of the zona matrix, we have established tissue culture lines that express each of the zona proteins and have developed transgenic mice that either lack a particular zona protein or express the human homologue. Breeding studies and in vitro sperm-egg binding studies are being used to investigate the role of individual zona proteins int he assemblage of the zona matrix and to determine the molecular basis for the specificity of fertilization.